As a research scientist in a pharmaceutical company, you are developing a new humancytokine (protein) therapeutic. This
Question:
As a research scientist in a pharmaceutical company, you are developing a new humancytokine (protein) therapeutic. This protein, ‘Glycoprotein X’, has been produced as arecombinant His6-tagged glycoprotein in Chinese Hamster Ovary (CHO) cell line culture. Other information that you have been provided with is that the native Glycoprotein X has a relative molecular weight of 35-39 kilodaltons, a pI of 4.2-4.5 and has three N-glycans and one O-glycan. You have been asked to purify sufficient quantities of the recombinant glycoprotein for protein structure determination by x-ray crystallography and for biochemical tests on its function.
a) Describe the workflow that you Design a PCR-based experiment to quantitatively measure the mRNA levels of the tumor suppressor proteins p53 and pRb plus the two oncogenes c-myc and K-ras in human cancer cells, in essay formatuse to purify recombinant Glycoprotein X and to check its purity. Provide information on the protein separation method(s) that you would select, how these method(s) work, and your expected end result. (70 marks)\\\\\\\\
b) After bioseparation, you find that Glycoprotein X still contains contaminating proteins. Based on the additional information that you have been provided with on the physicochemical properties of Glycoprotein X, briefly describe two additional‘polishing’ steps you could use to remove the remaining protein contaminants from your preparation of Glycoprotein X? (30 marks)
Financial Accounting an introduction to concepts, methods and uses
ISBN: 978-0324789003
13th Edition
Authors: Clyde P. Stickney, Roman L. Weil, Katherine Schipper, Jennifer Francis